Comparisons of mass spectrometry compatible surfactants for global analysis of the mammalian brain proteome.
Methods for the global analysis of protein expression offer an approach to study the molecular basis of disease. Studies of protein expression in tissue, such as brain, are complicated by the need for efficient and unbiased digestion of proteins that permit identification of peptides by shotgun proteomic methods. In particular, identification and characterization of less abundant membrane proteins has been of great interest for studies of brain physiology, but often proteins of interest are of low abundance or exist in multiple isoforms. Parsing protein isoforms as a function of disease will be essential. In this study, we develop a digestion scheme using detergents compatible with mass spectrometry that improves membrane protein identification from brain tissue. We show the modified procedure yields close to 5,000 protein identifications from 1.8 mg of rat brain homogenate with an average of 25% protein sequence coverage. This procedure achieves a remarkable reduction in the amount of starting material required to observe a broad spectrum of membrane proteins. Among the proteins identified from a mammalian brain homogenate, 1897 (35%) proteins are annotated by Gene Ontology as membrane proteins, and 1225 (22.6%) proteins are predicted to contain at least one transmembrane domain. Membrane proteins identified included neurotransmitter receptors and ion channels implicated in important physiological functions and disease.
Chen
Emily I
EI
Department of Chemical Physiology, The Scripps Research Institute, California 92037, USA.
McClatchy
Daniel
D
Park
Sung Kyu
SK
Yates
John R
JR
D000818:Animals; D001921:Brain / Q000166:cytology / Q000378:metabolism*; D057927:Hydrophobic and Hydrophilic Interactions; D013058:Mass Spectrometry / Q000379:methods*; D008565:Membrane Proteins / Q000032:analysis / Q000737:chemistry / Q000378:metabolism; D020543:Proteome / Q000032:analysis*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013501:Surface-Active Agents; D014357:Trypsin / Q000378:metabolism
D008565:Membrane Proteins; D020543:Proteome; D013501:Surface-Active Agents; D014357:Trypsin